Protein Expression of TLR2, TLR4, and TLR9 on Monocytes in TB, HIV, and TB/HIV.

Toll-like receptors (TLRs) have a critical role in recognizing pathogenic patterns and initiating immune responses against TB and HIV. Previously, studies described the gene expression of TLRs in patients with TB and HIV. Here, we demonstrated TLRs protein expressions and their association with clinical status and plasma markers in TB, HIV, and TB/HIV coinfection. The phenotyping of TLR2, TLR4, and TLR9 on CD14+ monocytes and their subsets were determined by multicolor flow cytometry. Host plasma biomarkers and microbial indices were measured using Luminex Multiplex assay and standard of care tools, respectively. TLR2 expression significantly enhanced in TB, slightly increased in HIV but slightly reduced in TB/HIV coinfection compared to apparently health controls (HC). On the other hand, TLR4 expression was significantly increased in TB, HIV, and TB/HIV compared to HC. Expression of TLR4 was equally enhanced on classical and intermediate monocytes while higher TLR2 expression on intermediate than classical monocytes. TLR4 had a positive correlation pattern with plasma biomarkers while TLR2 had an inverse correlation pattern. TLR4 is associated with disease severity while TLR2 is with the immune-competent status of patients. Our findings demonstrated that the pattern of TLR expression is disease as well as monocyte subset specific and distinct factors drive these differences.

Rare Variable M. tuberculosis Antigens induce predominant Th17 responses in human infection.

CD4 T cells are essential for immunity to M. tuberculosis (Mtb), and emerging evidence indicates that IL-17-producing Th17 cells contribute to immunity to Mtb. While identifying protective T cell effector functions is important for TB vaccine design, T cell antigen specificity is also likely to be important. To identify antigens that induce protective immunity, we reasoned that as in other pathogens, effective immune recognition drives sequence diversity in individual Mtb antigens. We previously identified Mtb genes under evolutionary diversifying selection pressure whose products we term Rare Variable Mtb Antigens (RVMA). Here, in two distinct human cohorts with recent exposure to TB, we found that RVMA preferentially induce CD4 T cells that express RoRγt and produce IL-17, in contrast to 'classical' Mtb antigens that induce T cells that produce IFNγ. Our results suggest that RVMA can be valuable antigens in vaccines for those already infected with Mtb to amplify existing antigen-specific Th17 responses to prevent TB disease.

Clinical and Imaging Characteristics of Smear Negative Pulmonary Tuberculosis Patients: A Comparative Study.

Background: Prevalence surveys in Ethiopia indicate smear negative pulmonary tuberculosis (SNPTB) taking the major share of the overall TB burden. It has also been a diagnostic dilemma worldwide leading to diagnostic delays and difficulty in monitoring treatment outcomes. This study determines and compares the clinical and imaging findings in SNPTB and smear positive PTB (SPPTB). Methodology. A case-control study was conducted on 313 PTB (173 SNPTB) patients. Data and sputum samples were collected from consented patients. Smear microscopy, GeneXpert, and culture analyses were performed on sputum samples. Data were analyzed using Stata version 17; a P value < 0.05 was considered statistically significant. Results: Of the 173 SNPTB patients, 42% were culture positive with discordances between test results reported by health facilities and Armauer Hansen Research Institute laboratory using concentrated smear microscopy. A previous history of TB and fewer cavitary lesions were significantly associated with SNPTB. Conclusions: Though overall clinical presentations of SNPTB patients resemble those seen in SPPTB patients, a prior history of TB was strongly associated with SNPTB. Subject to further investigations, the relatively higher discrepancies seen in TB diagnoses reflect the posed diagnostic challenges in SNPTB patients, as a higher proportion of these patients are also seen in Ethiopia.

Differential expression of chemokine receptors on monocytes in TB and HIV S.

In the present study, we defined multiple chemokine receptors expressed by classical, intermediate and non-classical monocyte subsets in TB, HIV and TB/HIV co-infection and associate it with the perturbation of monocyte subsets due to the diseases. Peripheral blood mononuclear cells from TB+ (n = 34), HIV+ (n = 35), TB + HIV+ (n = 12), as well as TB-HIV- healthy controls (n = 39), were tested for monocyte phenotyping by flow cytometry. Frequencies of intermediate and non-classical monocytes were significantly higher in TB and/or HIV disease relative to healthy controls. CCR2 and CX3CR1 were significantly higher on monocytes in TB disease, whereas CCR4 and CCR5 were present at higher levels in HIV disease. TB/HIV co-infected patients exhibited CCR2, CCR5 and CX3CR1 levels intermediate to TB and HIV subjects, while CCR4 was at a higher level than HIV. Despite the increase in the expression of chemokine receptors due to disease conditions, chemokine receptors maintained their original expression pattern on monocyte subsets. Our data provided new insight into the disease-specific but not monocyte subsets-specific modulation of chemokine receptors in TB and HIV.

Microbiological Investigations of Fine Needle Aspirates from Newly Suspected and Previously Treated Tubercular Lymphadenitis Patients.

Background: Extrapulmonary tuberculosis (EPTB), particularly tubercular lymphadenitis (TBLN), remains to pose a huge public health problem in Ethiopia. A significant number of TBLN patients who completed a full course anti-TB treatment regimen were reported to have enlarged lymph nodes and other TB-like clinical presentations. This could either be from a paradoxical reaction or microbiological relapse, possibly due to mono/multi-drug resistance. Objective: To investigate the rate of mono and multidrug resistance patterns of Mycobacterium tuberculosis as a cause of the observed treatment failures in clinically diagnosed and anti-TB treatment (newly or previously)-initiated LN patients. Methods: A cross-sectional study was conducted on 126 TBLN-suspected and previously treated patients between March and September 2022. Data were analyzed using SPSS (Version 26.0). Descriptive statistics were used to determine the frequency, percentage, sensitivity, specificity, and positive and negative predictive values. The level of agreement was determined using Cohen's kappa and a Chi-square test was used to measure the association between risk factors and laboratory test outcomes. A P-value <0.05 was considered statistically significant. Results: Mycobacterium tuberculosis was confirmed in 28.6% (N=36) of the 126 cases using BACTEC MGIT 960 culture detection method. Approximately, 13% (N=16) of the samples were collected from previously treated TBLN patients, of which 5/16 (31.3%) were multi-drug resistant, 7/16 were drug-sensitive and 4/16 were culture negative. To rule out other non-tuberculous agents, all samples were grown on blood and Mycosel agar plates, and no growth was detected. Conclusion: The emergence of drug resistant (DR) TB seems to not just be limited to pulmonary form but also to TBLN. In this study we observed a considerable number of microbiologically confirmed relapses among previously treated cases, possibly indicating the need for confirmation of drug resistance using rapid molecular methods or phenotypical methods during treatment follow up.

Diabetes mellitus is associated with an increased risk of unsuccessful treatment outcomes among drug-susceptible tuberculosis patients in Ethiopia: A prospective health facility-based study.

Background: The impact of diabetes mellitus on tuberculosis (TB) treatment outcomes has not been well investigated in most sub-Saharan countries including Ethiopia. The current study aimed to determine the association between diabetes mellitus and unsuccessful TB treatment outcomes among drug-susceptible TB patients treated at selected health facilities in Addis Ababa, Ethiopia. Methods: This health facility-based prospective cohort study was conducted at six randomly selected public health centers in Addis Ababa, from August 2020 until November 2021. Clinically diagnosed adult pulmonary and extra pulmonary TB patients were recruited at the time of treatment initiation. A multivariable logistic regression analysis was used to estimate the association between diabetes and unsuccessful TB treatment outcomes. Results: Among the total 267 enrolled participants, 9.7% of patients with TB were identified to have diabetes comorbidity. Of patients with diabetes and TB, 9 (34.6%) were newly diagnosed based on glucose test results. Despite an overall high TB treatment success rate (94.0%), more than one-fourth (26.9%) of patients with diabetes had a poor TB treatment outcome (26.9%), which was remarkably higher compared to patients without diabetes (3.7%). In multivariable regression, the adjusted odds of poor TB treatment outcome among those with diabetes was 14.8 (95% CI 3.5 - 62.7) times the odds of poor outcome patients without diabetes. Conclusion: Diabetes was significantly associated with increased odds of poor TB treatment outcomes among patients in Addis Ababa, Ethiopia.

Intravenous Immunoglobulin G (IVIG) Need Assessment Survey Toward Local Manufacturing of IVIG Using a Mini-Pool Plasma Fractionation Technique.

Immunoglobulin therapy has a crucial role in the treatment of primary and secondary immunodeficiencies as well as in a multitude of neurologic, hematologic, infectious, and autoimmune conditions. In the current study, a preliminary pilot scale needs assessment survey was conducted to examine the need for IVIG among patients in Addis Ababa, Ethiopia, and in so doing justify local manufacturing of IVIG products. The survey was performed by administering a structured questionnaire to private and government hospitals, a national blood bank, a regulatory body, and healthcare researchers working in academia and pharmaceutical companies. The questionnaire encompassed demographics and specific IVIG-related questions designed for each institution. Responses supplied in the study provide qualitative data. Our findings indicated that IVIG has been registered by the regulatory body for use in Ethiopia and there is a demand for the product in the country. The study also highlights that patients go as far as to clandestine markets to procure IVIG products at a cheaper price. To impede such illegal routes and make the product readily accessible, a small-scale and low-cost approach such as a mini-pool plasma fractionation technique could be implemented to locally purify and prepare IVIG using plasma collected through the national blood donation program.

Changes in plasma levels of endocrine hormones in lepromatous leprosy patients.

Background: Leprosy affects various endocrine glands and causes disorders in internal organs in addition to the skin and peripheral nerves. These disorders are often silent and remain undiagnosed or underreported. In particular, patterns of hormone changes during leprosy, especially in lepromatous leprosy (LL) patients, are often associated with dysregulation of different endocrine and sex hormones. The aim of this study was to assess changes in four endocrine hormones - namely cortisol, dehydroepiandrosterone (DHEA), growth hormone (GH), and leptin - among LL patients compared with apparently healthy controls. Method: In total, 80 plasma samples were systematically retrieved from a biorepository at the Armauer Hansen Research Institute (AHRI), based on quality, adequacy of sample volume, and appropriateness of linked clinical and sociodemographic data. Forty of the samples were obtained from LL patients (cases) and the remaining 40 from apparently healthy controls. Enzyme-linked immunosorbant assay (ELISA) was used to quantify levels of DHEA, cortisol, GH, and leptin hormones in the plasma samples. Data were analyzed using non-parametric statistics and the Mann-Whitney U-test (GraphPad Prism version 7.01). A p-value < 0.05 was considered statistically significant. Results: Plasma levels of cortisol concentration were significantly higher in LL cases (median = 111.4 ng/ml, range = 20.54-525.7) compared with healthy controls (median = 51.98 ng/ml, range = 3.805-328.4) (p = 0.003). Levels of GH and leptin were significantly lower in LL cases compared with healthy controls (median values for GH = 1.01 µIU/ml, range = 0.4625-86.82 and 2 µIU/ml, range = 0.5838-63.36, respectively (p = 0.022); median values for leptin = 891 pg/ml, range = 728.4-21816 and 5147 pg/ml, range = 730.4-52747, respectively (p < 0.0001)). There was an apparent reduction in the plasma levels of DHEA among LL cases compared with healthy controls (p = 0.297), although this difference was not statistically significant. Conclusion: Alterations in levels of endocrine hormones seen in LL patients reflect clinical and immunological conditions during lepromatous leprosy. However, large-scale studies are warranted to determine how leprosy causes such alterations in hormones and the interplay between endocrine hormones and the immune system during leprosy disease.

The Impact of Diabetes and Prediabetes on Prevalence of Mycobacterium tuberculosis Infection Among Household Contacts of Active Tuberculosis Cases in Ethiopia.

Background: It is uncertain whether diabetes affects the risk of developing latent tuberculosis infection (LTBI) following exposure to Mycobacterium tuberculosis (Mtb). We assessed the relationship of diabetes or prediabetes and LTBI among close and household contacts (HHCs) of patients with active pulmonary tuberculosis (TB) disease in Addis Ababa, Ethiopia. Methods: In this cross-sectional study, we performed interferon-γ release assays, TB symptom screening, and point-of-care glycolated hemoglobin (HbA1c) testing among HHCs of active TB cases. Diabetes status was classified into diabetes (HbA1c ≥6.5% or self-reported diagnosis), prediabetes (5.7%-6.4%), and euglycemia (≤5.6%). Multivariable logistic regression was used to determine the association of diabetes with LTBI. Results: Among 597 study participants, 123 (21%) had dysglycemia including diabetes (n = 31) or prediabetes (n = 92); 423 (71%) participants were diagnosed with LTBI. Twelve of 31 (39%) HHCs with diabetes were previously undiagnosed with diabetes. The prevalence of LTBI among HHCs with diabetes, prediabetes, and euglycemia was 87% (27/31), 73% (67/92), and 69% (329/474), respectively. In multivariable analysis adjusted for age, sex, and HIV status, the odds of LTBI among HHCs with diabetes were 2.33 (95% confidence interval [CI], .76-7.08) times the odds of LTBI without diabetes. When assessing interaction with age, the association of diabetes and LTBI was robust among participants aged ≥40 years (adjusted odds ratio [aOR], 3.68 [95% CI, .77-17.6]) but not those <40 years (aOR, 1.15 [95% CI, .22-6.1]). Conclusions: HHCs with diabetes may be more likely to have LTBI than those with euglycemia. Further investigations are needed to assess mechanisms by which diabetes may increase risk of LTBI after Mtb exposure.

Evaluation of the diagnostic performance of PanbioTM Abbott SARS-CoV-2 rapid antigen test for the detection of COVID-19 from suspects attending ALERT center.

BACKGROUND: The emergence and rapid spread of coronavirus disease 2019 (COVID-19), a potentially lethal disease caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), is causing public health issues around the world. In resource-constrained nations, rapid Abbott SARS-CoV-2 antigen test kits are critical for addressing diagnostic gaps in health institutions and community screening. However, there is no evidence or proof of diagnostic performance in Ethiopia. The aim of this study was to compare the performance of PanbioTM Abbott SARS-CoV-2antigen rapid test kit to the gold standard, RT-PCR, in COVID-19 patients with clinical symptoms suggestive of COVID-19. METHOD: A prospective, cross-sectional study was conducted between November 2021 and April 2022, on 120 suspected patients recruited from outpatient, emergency, and intensive care units in one of the tertiary hospitals in Ethiopia. Nasopharyngeal swabs were collected from suspected cases and were tested using the Abbott SARS-CoV-2 kit, a rapid diagnostic test (RDT) and compared to the reference standard RT-PCR. RESULT: The sensitivity and specificity of the RDT were 74.2% and 100%, respectively. A total of 62 samples (51.6%) were RT-PCR positive. Of these, 46 were Ag-RDT positive. Sensitivity among symptomatic patients was 79.4% (95% CI 68.3-90). The Abbot RDT and RT-PCR had a Kappa value of agreement of 0.735 (p < 0.001). These values were acceptable when compared to the WHO's suggested thresholds. CONCLUSION: The finding from this study support the use of the Abbot RDT as a diagnostic tool in COVID-19 suspects, mainly in those with higher viral loads.

Persistent expression of activation markers on Mycobacterium tuberculosis-specific CD4 T cells in smear negative TB patients.

BACKGROUND: T cell activation (HLA-DR, CD-38), proliferation (KI-67), and functional (IFN-γ, TNF-α) markers have recently been shown to be useful in predicting and monitoring anti-TB responses in smear positive TB, but previous research did not characterize the activation and proliferation profiles after therapy of smear negative TB. METHODOLOGY: In this study, we used polychromatic flow cytometry to assess selected PPD-specific T cell markers using fresh PBMC of smear negative and positive pulmonary tuberculosis (PTB) patients, recruited from health facilities in Addis Ababa. RESULT: Levels of activation (HLA-DR, CD38) and proliferation (Ki-67) among total unstimulated CD4 T cells decreased significantly after therapy, particularly at month 6. Similarly, levels of PPD-specific T cell activation markers (HLA-DR, CD-38) were significantly lower in smear positive PTB patients following treatment, whereas a consistent decline in these markers was less apparent among smear negative PTB patients at the sixth month. CONCLUSION: After six months of standard anti-TB therapy, persistent levels of activation of HLA-DR and CD-38 from PPD specific CD4+T cells in this study could indicate that those markers have little value in monitoring and predicting anti-TB treatment response in smear negative pulmonary TB patients in Ethiopian context.

A Single Dose of ChAdOx1 nCoV-19 Vaccine Elicits High Antibody Responses in Individuals with Prior SARS-CoV-2 Infection Comparable to That of Two-Dose-Vaccinated, SARS-CoV-2-Infection-Naïve Individuals: A Longitudinal Study in Ethiopian Health Workers.

Single-dose COVID-19 vaccines, mostly mRNA-based vaccines, are shown to induce robust antibody responses in individuals who were previously infected with SARS-CoV-2, suggesting the sufficiency of a single dose for those individuals in countries with limited vaccine supply. However, these important data are limited to developed nations. We conducted a prospective longitudinal study among Ethiopian healthcare workers who received a ChAdOx1 nCoV-19 vaccine. We compared the geometric mean titers (GMTs) of the SARS-CoV-2 receptor-binding domain (RBD)-specific IgG antibodies in 39 SARS-CoV-2 naïve participants and 24 participants previously infected with SARS-CoV-2 (P.I.), who received two doses of ChAdOx1 nCoV-19 vaccine across the two post-vaccination time points (at 8 to 12 weeks post single dose and two dose vaccinations). We noted that the GMT (1632.16) in naïve participants at 8-12 weeks post first dose were comparable to the GMT (1674.94) observed in P.I. participants prior to vaccination. Interestingly, P.I. participants had significantly higher antibody titers compared to naïve participants, after both the first (GMT, 4913.50 vs. 1632.16) and second doses (GMT, 9804.60 vs. 6607.30). Taken together, our findings show that a single ChAdOx1 nCoV-19 dose in previously SARS-CoV-2 infected individuals elicits similar, if not higher, antibody responses to those of two-dose-vaccinated naïve individuals.

Mycobacterium tuberculosis antigen-specific T-cell responses in smear-negative pulmonary tuberculosis patients.

Despite recent improvements in microbial detection, smear-negative TB remains a diagnostic challenge. In this study, we investigated the potential discriminatory role of polychromatic flow cytometry of M. tuberculosis antigen-specific T cells to discriminate smear-negative TB from health controls with or without latent TB infection, and non-TB respiratory illnesses in an endemic setting. A cross-sectional study was conducted on HIV negative, newly diagnosed smear-positive PTB (n = 34), smear-negative/GeneXpert negative PTB (n = 29) patients, non-TB patients with respiratory illness (n = 33) and apparently healthy latent TB infected (n = 30) or non-infected (n = 23) individuals. The expression of activation (HLA-DR, CD-38), proliferation (Ki-67), and functional (IFN-γ, TNF-α) T-cell markers using polychromatic flow cytometry was defined after stimulation with PPD antigens. Sputum samples were collected and processed from all patients for Mtb detection using a concentrated microscopy, LJ/MGIT culture, and RD9 typing by PCR. Our study showed CD4 T cells specific for PPD co-expressed activation/proliferation markers together with induced cytokines IFN-γ or TNF-α were present at substantially higher levels among patients with smear-positive and smear-negative pulmonary TB than among healthy controls and to a lesser extent among patients with non-TB illness. Our study conclude that smear-negative TB can be distinguished from non-TB respiratory illness and healthy controls with a flow cytometric assay for PPD-specific T cells co-expressing activation/proliferation markers and cytokines.

High seroprevalence of anti-SARS-CoV-2 antibodies among Ethiopian healthcare workers.

BACKGROUND: COVID-19 pandemic has a devastating impact on the economies and health care system of sub-Saharan Africa. Healthcare workers (HWs), the main actors of the health system, are at higher risk because of their occupation. Serology-based estimates of SARS-CoV-2 infection among HWs represent a measure of HWs' exposure to the virus and could be used as a guide to the prevalence of SARS-CoV-2 in the community and valuable in combating COVID-19. This information is currently lacking in Ethiopia and other African countries. This study aimed to develop an in-house antibody testing assay, assess the prevalence of SARS-CoV-2 antibodies among Ethiopian high-risk frontline HWs. METHODS: We developed and validated an in-house Enzyme-Linked Immunosorbent Assay (ELISA) for specific detection of anti-SARS-CoV-2 receptor binding domain immunoglobin G (IgG) antibodies. We then used this assay to assess the seroprevalence among HWs in five public hospitals located in different geographic regions of Ethiopia. From consenting HWs, blood samples were collected between December 2020 and February 2021, the period between the two peaks of COVID-19 in Ethiopia. Socio-demographic and clinical data were collected using questionnaire-based interviews. Descriptive statistics and bivariate and multivariate logistic regression were used to determine the overall and post-stratified seroprevalence and the association between seropositivity and potential risk factors. RESULTS: Our successfully developed in-house assay sensitivity was 100% in serum samples collected 2- weeks after the first onset of symptoms whereas its specificity in pre-COVID-19 pandemic sera was 97.7%. Using this assay, we analyzed a total of 1997 sera collected from HWs. Of 1997 HWs who provided a blood sample, and demographic and clinical data, 51.7% were females, 74.0% had no symptoms compatible with COVID-19, and 29.0% had a history of contact with suspected or confirmed patients with SARS-CoV-2 infection. The overall seroprevalence was 39.6%. The lowest (24.5%) and the highest (48.0%) seroprevalence rates were found in Hiwot Fana Specialized Hospital in Harar and ALERT Hospital in Addis Ababa, respectively. Of the 821 seropositive HWs, 224(27.3%) of them had a history of symptoms consistent with COVID-19 while 436 (> 53%) of them had no contact with COVID-19 cases as well as no history of COVID-19 like symptoms. A history of close contact with suspected/confirmed COVID-19 cases is associated with seropositivity (Adjusted Odds Ratio (AOR) = 1.4, 95% CI 1.1-1.8; p = 0.015). CONCLUSION: High SARS-CoV-2 seroprevalence levels were observed in the five Ethiopian hospitals. These findings highlight the significant burden of asymptomatic infection in Ethiopia and may reflect the scale of transmission in the general population.

Integration of cytopathology with molecular tests to improve the lab diagnosis for TBLN suspected patients.

BACKGROUND: Tuberculosis lymphadenitis (TBLN) diagnosis is often challenging in most resource poor settings. Often cytopathologic diagnosis of TBLN suspected patients is inconclusive impeding timely clinical management of TBLN suspected patients, further exposing suspected patients either for unnecessary use of antibiotics or empirical treatment. This may lead to inappropriate treatment outcome or more suffering of suspected patients from the disease. In this study, an integrated diagnostic approach has been evaluated to elucidate its utility in the identification of TBLN suspected patients. METHODS: A cross-sectional study was conducted on 96 clinically diagnosed TBLN suspected patients, where fine needle aspirate (FNA) samples were collected at the time of diagnosis. FNA cytology, Ziehl-Neelsen (ZN), Auramine O (AO) staining, GeneXpert MTB/RIF and Real time PCR (RT-PCR) were performed on concentrated FNA samples. Considering culture as a gold standard, the sensitivity, specificity, positive and negative predictive values were calculated. Cohen's Kappa value was used to measure interrater variability and level of agreement and a P-value of <0.05 was considered as statistically significant. RESULT: Out of the 96 FNA sample, 12 (12.5%) were identified to have Mycobacterium tuberculosis (Mtb) using ZN staining, 27 (28.1%) using AO staining, 51 (53.2%) using FNAC, 43 (44.7%) using GeneXpert MTB/RIF, 51 (53.1%) using Real time PCR (RT-PCR) and 36 (37.5%) using Lowenstein-Jensen (LJ) culture. Compared to LJ culture, the sensitivities of GeneXpert MTB/RIF, RT-PCR, and FNAC were 91.7%, 97.2%, and 97.2%, respectively and the specificities were 83.3%, 73.3%, and 68.3%, respectively. GeneXpert MTB/RIF and RT-PCR when combined with FNAC detected 61 (63.5%) cases as having Mtb, and the sensitivity and specificity was 100% and 58.3%, respectively. CONCLUSION: FNA cytology and RT-PCR detected more TBLN cases compared to other Mtb detection tools and the detection sensitivity even improved when FNA cytology was combined with GeneXpert MTB/RIF, performed on concentrated FNA sample, suggesting the combined tests as an alternative approach for improved diagnosis of TBLN.

Alteration of Endocrine Hormones and Antibody Responses in Different Spectrum of Tuberculosis Disease.

Effective control of Mycobacterium tuberculosis (Mtb) infection is mediated by multifaceted factors that involve both the endocrine and immune system. Profiling hormones and antibodies in different stages of TB provides insight in the pathogenesis of the disease. In this study, we profiled endocrine hormones (dehydroepiandrosterone (DHEA), cortisol, testosterone, estradiol, growth hormone and leptins) and Mtb strain H37RV lipoarabinomannan (LAM)-specific antibody levels in plasma samples, collected from pulmonary TB (PTB) patients, TB lymphadenitis (TBLN) patients and latently infected (QFT-positive) or uninfected (QFT-negative) apparently healthy individuals using ELISA. Plasma levels of leptin and DHEA were significantly low in PTB and TBLN patients compared to healthy controls (P<0.0001 and P=0.02, respectively), whereas these levels significantly increased following anti-TB treatment (P=0.002 and P=0.0001, respectively) among TB patients. The levels of estradiol and testosterone significantly improved following anti-TB treatment (P=0.03 and P=0.0003, respectively), whereas cortisol and growth hormones declined significantly (P <0.05). Similarly, LAM-specific IgG, IgM and IgA were significantly higher in PTB patients compared to other groups, whereas levels of IgG1 subtype were significantly higher among LTBI groups compared to both TB patients and QFT-negative individuals (P<0.0001). Overall, we observed significantly variable levels of endocrine hormones as well as immunoglobulins across the spectrum of TB illness and such profiling has a significant contribution in selection of effective biomarkers that have roles in TB treatment monitoring or diagnostics. Although this study did not show a functional association between hormones and antibodies, alterations in the levels of these biomarkers suggest the key roles these markers play in TB pathogenesis.

A single dose ChAdOx1 nCoV-19 vaccine elicits high antibody responses in individuals with prior SARS-CoV-2 infection comparable to that of double dose vaccinated SARS-CoV-2 infection naïve individuals.

Background A single dose COVID-19 vaccines, mostly mRNA-based vaccines, are shown to induce robust antibody responses in individuals who were previously infected with SARS-CoV-2, suggesting the sufficiency of a single dose to those individuals. However, these important data are limited to developed nations and lacking in resource-limited countries, like Ethiopia. Methods We compared receptor-binding domain (RBD)-specific IgG antibodies in 40 SARS-CoV-2 naïve participants and 25 participants previously infected with SARS-CoV-2, who received two doses of ChAdOx1 nCoV-19 vaccine. We measured the antibody response in post-vaccination blood samples from both groups of participants collected at four different post-vaccination time points: 8- and 12-weeks after each dose of the vaccine administration using an in-house developed ELISA. Results We observed a high level of anti-RBD IgG antibodies titers 8-weeks after a single dose administration (16/27; 59.3%) among naïve participants, albeit dropped significantly (p<0.05) two months later, suggesting the protective immunity elicited by the first dose ChAdOx1 nCoV-19 vaccine will likely last for a minimum of three months. However, as expected, a significant (p<0.001) increase in the level of anti-RBD IgG antibodies titers was observed after the second dose administration in all naïve participants. By contrast, the ChAdOx1 nCoV-19 vaccine-induced anti-RBD IgG antibody titers produced by the P.I participants at 8- to 12-weeks post-single dose vaccination were found to be similar to the antibody titers seen after a two-dose vaccination course among infection- naïve participants and showed no significant (p>0.05) increment following the second dose administration. Conclusion Taken together, our findings show that a single ChAdOx1 nCoV-19 dose in previously SARS-CoV-2 infected individuals elicits similar antibody responses to that of double dose vaccinated naïve individuals. Age and sex were not associated with the level of vaccine-elicited immune responses in both individuals with and without prior SARS-CoV-2 infection. Further studies are required to assess the need for a booster dose to extend the duration and amplitude of the specific protective immune response in Ethiopia settings, especially following the Omicron pandemic.

Saliva is superior over nasopharyngeal swab for detecting SARS-CoV2 in COVID-19 patients.

Scaling up of diagnostic capacity is needed to mitigate the global pandemic of SARS-CoV2. However, there are challenges including shortage of sample collection swabs and transport medium. Saliva has been recommended as a simple, low-cost, non-invasive option. However, data from different populations and settings are limited. Here, we showed that saliva could be a good alternative sample to diagnose COVID-19 patients. Pair of NPS-saliva samples was collected from 152 symptomatic; confirmed COVID-19 patients, and compared their positivity rate, viral load, and duration of viral shedding. From 152 patients, 80 (52.63%) tested positive and 72 (47.37%) were negative for SARSA-CoV2 in NPS sample. In saliva, 129 (92.14%) were tested positive and 11 (7.86%) were negative on the day of admission to hospital. The overall percent agreement of RT-PCR result of Saliva to NPS was 70% (196/280). A comparison of viral load from 72 NPS-saliva pair samples on day of admission shows saliva contains significantly higher viral load (P < 0.001). In conclusion, saliva has higher yield in detecting SARS-CoV2, and COVID-19 patients show higher viral load and prolonged period of viral shedding in saliva. Therefore, we recommend saliva as a better alternative sample to NPS to diagnose COVID-19 patients.

PDL1 expression on monocytes is associated with plasma cytokines in Tuberculosis and HIV.

INTRODUCTION: PDL1 and its interaction with PD1 is implicated in immune dysfunction in TB and HIV. The expression of PDL1 on multiple subsets of monocytes as well as their associations with cytokines and microbial products have not been well studied. METHOD: HIV (TB-HIV+), TB (TB+HIV-) and TB/HIV co-infected (TB+HIV+) patients as well as apparently healthy controls (TB-HIV-) were recruited. TB and HIV patients were treatment naïve while TB/HIV patients were both ART naïve and experienced but not yet started TB therapy. Monocyte subsets were evaluated for PDL1 expression by flow cytometry; plasma TNFα, IL6, IP10, IFNγ and IL10 were measured by Luminex; and cytokine mRNA from purified monocytes quantitated by qPCR. The association of PDL1 with cytokines, clinical and microbial indices, including HIV viral load, TB smear microscopy and TB urinary lipoarabinomannan (LAM) were assessed. RESULTS: Monocyte expression of PDL1 was significantly higher in TB, HIV and TB/HIV co-infected patients compared with healthy controls (p = 0.0001), with the highest levels in TB/HIV co-infected patients. The highest expression of PDL1 was on intermediate (CD14+CD16+) monocytes in all participant groups. PDL1 strongly correlated with HIV viral load in TB/HIV while weakly correlated in HIV. PDL1 levels moderately correlated with plasma TNFα, IL6, IP10, IFNγ and IL10 level in TB subjects whereas weakly correlated with TNFα and IP10 in HIV patients. However, cytokine mRNA from purified monocytes showed no association with either plasma cytokines or monocyte PDL1 expression, implying that if cytokines modulate PDL1, they are likely not originating from circulating monocytes themselves. These results underscore the importance of further characterization of multiple monocyte subsets and their phenotypic and functional differences in different disease states.